Genomic characteristics of nine Nitrospirota metagenome-assembled genomes in deep-sea sediments from East Pacific polymetallic nodules zone.

Previously studies have reported that MAGs (Metagenome-assembled genomes) belong to "Candidatus Manganitrophaceae" of phylum Nitrospirota with chemolithoautotrophic manganese oxidation potential exist in freshwater and hydrothermal environments. However, Nitrospirota members with chemolithoautotrophic manganese oxidation potential have not been reported in other marine environments. Through metagenomic sequencing, assembly and binning, nine metagenome-assembled genomes belonging to Nitrospirota are recovered from sediment of different depths in the polymetallic nodule area. Through the key functional genes annotation results, we find that these Nitrospirota have limited potential to oxidize organic carbon because of incomplete tricarboxylic acid cycle and most of them (6/9) have carbon dioxide fixation potential through different pathway (rTCA, WL or CBB). One MAG belongs to order Nitrospirales has the potential to use manganese oxidation to obtain energy for carbon fixation. In addition to manganese ions, the oxidation of inorganic nitrogen, sulfur, hydrogen and carbon monoxide may also provide energy for the growth of these Nitrospirota. In addition, different metal ion transport systems can help those Nitrospirota to resist heavy metal in sediment. Our work expands the understanding of the metabolic potential of Nitrospirota in sediment of polymetallic nodule region and may contributes to promoting the study of chemolithoautotrophic manganese oxidation.

mARC1 in MASLD: Modulation of lipid accumulation in human hepatocytes and adipocytes.

BACKGROUND: Mutations in the gene MTARC1 (mitochondrial amidoxime-reducing component 1) protect carriers from metabolic dysfunction-associated steatohepatitis (MASH) and cirrhosis. MTARC1 encodes the mARC1 enzyme, which is localized to the mitochondria and has no known MASH-relevant molecular function. Our studies aimed to expand on the published human genetic mARC1 data and to observe the molecular effects of mARC1 modulation in preclinical MASH models. METHODS AND RESULTS: We identified a novel human structural variant deletion in MTARC1, which is associated with various biomarkers of liver health, including alanine aminotransferase levels. Phenome-wide Mendelian Randomization analyses additionally identified novel putatively causal associations between MTARC1 expression, and esophageal varices and cardiorespiratory traits. We observed that protective MTARC1 variants decreased protein accumulation in in vitro overexpression systems and used genetic tools to study mARC1 depletion in relevant human and mouse systems. Hepatocyte mARC1 knockdown in murine MASH models reduced body weight, liver steatosis, oxidative stress, cell death, and fibrogenesis markers. mARC1 siRNA treatment and overexpression modulated lipid accumulation and cell death consistently in primary human hepatocytes, hepatocyte cell lines, and primary human adipocytes. mARC1 depletion affected the accumulation of distinct lipid species and the expression of inflammatory and mitochondrial pathway genes/proteins in both in vitro and in vivo models. CONCLUSIONS: Depleting hepatocyte mARC1 improved metabolic dysfunction-associated steatotic liver disease-related outcomes. Given the functional role of mARC1 in human adipocyte lipid accumulation, systemic targeting of mARC1 should be considered when designing mARC1 therapies. Our data point to plasma lipid biomarkers predictive of mARC1 abundance, such as Ceramide 22:1. We propose future areas of study to describe the precise molecular function of mARC1, including lipid trafficking and subcellular location within or around the mitochondria and endoplasmic reticulum.

Topological Structure Realized in Cove-Edged Graphene Nanoribbons via Incorporation of Periodic Pentagon Rings.

Cove-edged zigzag graphene nanoribbons are predicted to show metallic, topological, or trivial semiconducting band structures, which are precisely determined by their cove offset positions at both edges as well as the ribbon width. However, due to the challenge of introducing coves into zigzag-edged graphene nanoribbons, only a few cove-edged graphene nanoribbons with trivial semiconducting bandgaps have been realized experimentally. Here, we report that the topological band structure can be realized in cove-edged graphene nanoribbons by embedding periodic pentagon rings on the cove edges through on-surface synthesis. Upon noncontact atomic force microscopy and scanning tunneling spectroscopy measurements, the chemical and electronic structures of cove-edged graphene nanoribbons with periodic pentagon rings have been characterized for different lengths. Combined with theoretical calculations, we find that upon inducing periodic pentagon rings the cove-edged graphene nanoribbons exhibit nontrivial topological structures. Our results provide insights for the design and understanding of the topological character in cove-edged graphene nanoribbons.

TGN-020 Alleviate Inflammation and Apoptosis After Cerebral Ischemia-Reperfusion Injury in Mice Through Glymphatic and ERK1/2 Signaling Pathway.

Post-stroke acute inhibition of aquaporin 4 (AQP4) is known to exacerbate inflammation and apoptosis, yet the underlying mechanisms are not fully understood. The objective of this study was to investigate the specific mechanism of inflammation and apoptosis following cerebral ischemia-reperfusion (I/R) injury using the AQP4-specific inhibitor, N-(1,3,4-thiadiazol-2-yl) pyridine-3-carboxamide dihydrochloride (TGN-020). Ischemic stroke was induced in mice using the middle cerebral artery occlusion (MCAO) model. The C57/BL6 mice were randomly divided into three groups as follows: sham operation, I/R 48 h, and TGN-020 + I/R 48 h treatment. All mice were subjected to a series of procedures. These procedures encompassed 2,3,5-triphenyltetrazolium chloride (TTC) staining, neurological scoring, fluorescence tracing, western blotting, immunofluorescence staining, and RNA sequencing (RNA-seq). The glymphatic function in the cortex surrounding cerebral infarction was determined using tracer, glial fibrillary acid protein (GFAP), AQP4 co-staining, and beta-amyloid precursor protein (APP) staining; differential genes were detected using RNA-seq. The influence of TGN-020 on the extracellular signal-regulated kinase 1/2 (ERK) 1/2 pathway was confirmed using the ERK1/2 pathway agonists Ro 67-7467. Additionally, we examined the expression of inflammation associated with microglia and astrocytes after TGN-020 and Ro 67-7467 treatment. Compared with I/R group, TGN-020 alleviated glymphatic dysfunction by inhibiting astrocyte proliferation and reducing tracer accumulation in the peri-infarct area. RNA-seq showed that the differentially expressed genes were mainly involved in the activation of astrocytes and microglia and in the ERK1/2 pathway. Western blot and immunofluorescence further verified the expression of associated inflammation. The inflammation and cell apoptosis induced by I/R are mitigated by TGN-020. This mitigation occurs through the improvement of glymphatic function and the inhibition of the ERK1/2 pathway.

Limb Remote Ischemic Postconditioning Improves Glymphatic Dysfunction After Cerebral Ischemia-Reperfusion Injury.

BACKGROUND: Delayed neuronal damage can be caused or aggravated after cerebral ischemia-reperfusion (I/R) injury. Recent studies have shown that glymphatic system dysfunction after cerebral ischemia-reperfusion injury is involved in ischemic brain edema and neuroinflammation, thereby regulating cerebral ischemia-reperfusion injury. The aim of this study is to investigate the changes of glymphatic system after cerebral ischemia-reperfusion injury and whether limb remote ischemic postconditioning (LRIP) can improve the function of glymphatic system to protect the brain. METHODS: To establish a focal brain I/R injury mouse model, this study utilized the middle cerebral artery occlusion/reperfusion (MCAO/R) method. The present study classified eight-week-old C57BL/6 male mice into three groups. The changes in glymphatic function in different periods of ischemia and reperfusion were analyzed through immunofluorescence, transmission electron microscopy (TEM), and Western-Blot (WB) assays. The contents of the evaluation included cerebrospinal fluid flow, swelling degree of brain tissue, aquaporin-4 (AQP4) expression and polarization, and amyloid-ß (Aß) excretion. RESULTS: In the early stages of cerebral ischemia, cerebrospinal fluid (CSF) flow is disturbed, accompanied by a decrease in AQP4 polarization. The polarity of AQP4 decreased from 12 h to 72 h of reperfusion, the Aß deposition. LRIP can increase the expression of ß-DG and AQP4 polarization, reduce the deposition of Aß, improve the function of the glymphatic system, and reduce the expression of AQP4 to play A protective role in brain. CONCLUSION: Glymphatic system impaired after cerebral ischemia-reperfusion injury in mice. LRIP may play a neuroprotective role by improving glymphatic function after I/R.

Rubber band ligation versus coagulation for the treatment of haemorrhoids: a meta-analysis of randomised controlled trials.

Non-surgical therapies have the advantage of lower postoperative pain and complication rates compared with surgical therapies. Rubber band ligation and coagulation are two kinds of non-surgical therapies. The aim of this study is to compare the clinical outcomes of rubber band ligation and coagulation. A systematic review was conducted to identify randomised clinical trials that compare rubber band ligation and coagulation treatments for haemorrhoids. PubMed and Web of Science were searched, from inception to April 30th,2022. Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines were followed. Fifty-nine studies were identified. Nine trials met the inclusion criteria. All trials were of moderate methodological quality. No significant difference was found between rubber band ligation and coagulation in terms of efficacy rate, postoperative prolapse rate, recurrence rate and postoperative urine retention rate after treatment. Patients undergoing rubber band ligation had higher postoperative pain rate and lower postoperative bleeding rate than patients undergoing coagulation. The subgroup analysis showed that there was no significant difference between rubber band ligation and infrared coagulation or non-infrared coagulation in terms of efficacy rate, postoperative bleeding and postoperative urine retention rate after treatment. Patients undergoing rubber band ligation had a higher postoperative pain rate than patients undergoing infrared coagulation or non-infrared coagulation. We believe that coagulation for haemorrhoids still has a good future. PROSPERO registration number CRD42022311281.

FinnGen provides genetic insights from a well-phenotyped isolated population.

Population isolates such as those in Finland benefit genetic research because deleterious alleles are often concentrated on a small number of low-frequency variants (0.1% ≤ minor allele frequency < 5%). These variants survived the founding bottleneck rather than being distributed over a large number of ultrarare variants. Although this effect is well established in Mendelian genetics, its value in common disease genetics is less explored1,2. FinnGen aims to study the genome and national health register data of 500,000 Finnish individuals. Given the relatively high median age of participants (63 years) and the substantial fraction of hospital-based recruitment, FinnGen is enriched for disease end points. Here we analyse data from 224,737 participants from FinnGen and study 15 diseases that have previously been investigated in large genome-wide association studies (GWASs). We also include meta-analyses of biobank data from Estonia and the United Kingdom. We identified 30 new associations, primarily low-frequency variants, enriched in the Finnish population. A GWAS of 1,932 diseases also identified 2,733 genome-wide significant associations (893 phenome-wide significant (PWS), P < 2.6 × 10-11) at 2,496 (771 PWS) independent loci with 807 (247 PWS) end points. Among these, fine-mapping implicated 148 (73 PWS) coding variants associated with 83 (42 PWS) end points. Moreover, 91 (47 PWS) had an allele frequency of <5% in non-Finnish European individuals, of which 62 (32 PWS) were enriched by more than twofold in Finland. These findings demonstrate the power of bottlenecked populations to find entry points into the biology of common diseases through low-frequency, high impact variants.

A cancer stem cell associated gene signature for predicting overall survival of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the most prevalent type of primary liver cancer characterized by high mortality and morbidity rate. The lack of effective treatments and the high frequency of recurrence lead to poor prognosis of patients with HCC. Therefore, it is important to develop robust prediction tools for predicting the prognosis of HCC. Recent studies have shown that cancer stem cells (CSC) participate in HCC progression. The aim of this study was to explore the prognostic value of CSC-related genes and establish a prediction model based on data from The Cancer Genome Atlas (TCGA) database. In this study, 475 CSC-related genes were obtained from the Molecular Signature Database and 160 differentially expressed CSC-related genes in HCC patients were identified using the limma R package in the TCGA database. A total of 79 CSC-related genes were found to be associated with overall survival (OS). Using the least absolute shrinkage and selection operator (LASSO) and multivariate Cox regressions, a 3-gene signature (RAB10, TCOF1, and PSMD14) was constructed. Receiver operating characteristic (ROC) curves and Kaplan-Meier survival curves were constructed to test the prediction performance of the signature. Performance of the signature was validated using the International Cancer Genome Consortium (ICGC) dataset. In addition, immune feature and functional enrichment analyses were carried out to explore the underlying mechanisms. Moreover, a co-expression network was constructed using the weighted gene correlation network analysis (WGCNA) method to select genes significantly associated with risk scores in HCC in the TCGA dataset. The SGO2 gene was found to be significantly associated with risk scores of HCC. In vitro experiments revealed that it can promote HCC cell proliferation. Therefore, SGO2 may be a potential therapeutic target for HCC treatment. The constructed nomogram can help clinicians make decisions about HCC treatment.

NLK is required for Ras/ERK/SRF/ELK signaling to tune skeletal muscle development by phosphorylating SRF and antagonizing the SRF/MKL pathway.

Serum response factor (SRF) regulates differentiation and proliferation by binding to RhoA-actin-activated MKL or Ras-MAPK-activated ELK transcriptional coactivators, but the molecular mechanisms responsible for SRF regulation remain unclear. Here, we show that Nemo-like kinase (NLK) is required for the promotion of SRF/ELK signaling in human and mouse cells. NLK was found to interact with and phosphorylate SRF at serine residues 101/103, which in turn enhanced the association between SRF and ELK. The enhanced affinity of SRF/ELK antagonized the SRF/MKL pathway and inhibited mouse myoblast differentiation in vitro. In a skeletal muscle-specific Nlk conditional knockout mouse model, forming muscle myofibers underwent hypertrophic growth, resulting in an increased muscle and body mass phenotype. We propose that both phosphorylation of SRF by NLK and phosphorylation of ELKs by MAPK are required for RAS/ELK signaling, confirming the importance of this ancient pathway and identifying an important role for NLK in modulating muscle development in vivo.

Cyanide-free synthesis of aromatic nitriles from aldoximes: Discovery and application of a novel heme-containing aldoxime dehydratase.

Aromatic nitriles are important structural motifs that frequently existed in pharmaceutical drugs. Due to the convenient synthesis of aldoximes from aldehydes, the dehydration of aldoximes to corresponding nitriles by aldoxime dehydratases (Oxds) is considered as a safe and robust enzymatic production route. Although the Oxd genes are widely distributed in microbial kingdom, so far less than ten Oxds were expressed and further characterized. In this study, we found 26 predicted putative Oxd genes from the GenBank database using a genome mining strategy. The Oxd gene from Pseudomonas putida F1 was cloned and functionally expressed in Escherichia coli BL21 (DE3). The amino acid sequence of OxdF1 shows high identities of 33∼85 % to other characterized Oxds, and contained a ferrous heme as the catalytic site. The optimum reaction pH and temperature of recombinant OxdF1 were 7.0 and 35 °C, respectively. OxdF1 was stable in pH 7.0 potassium phosphate buffer at 30 °C, and its half-life was approximately 3.8 h. OxdF1 can efficiently dehydrate aromatic and heterocyclic aldoximes to nitriles, such as 2-bromobenzaldoxime, 2-chloro-6-fluorobenzaldoxime, thiophene-2-carboxaldoxime, and pyridine-3-aldoxime. Therefore, the recombinant OxdF1 shows a potential application in the cyanide-free synthesis of aromatic nitriles.

Intrinsic bioactivity of black phosphorus nanomaterials on mitotic centrosome destabilization through suppression of PLK1 kinase.

Although nanomaterials have shown promising biomedical application potential, incomplete understanding of their molecular interactions with biological systems prevents their inclusion into mainstream clinical applications. Here we show that black phosphorus (BP) nanomaterials directly affect the cell cycle's centrosome machinery. BP destabilizes mitotic centrosomes by attenuating the cohesion of pericentriolar material and consequently leads to centrosome fragmentation within mitosis. As a result, BP-treated cells exhibit multipolar spindles and mitotic delay, and ultimately undergo apoptosis. Mechanistically, BP compromises centrosome integrity by deactivating the centrosome kinase polo-like kinase 1 (PLK1). BP directly binds to PLK1, inducing its aggregation, decreasing its cytosolic mobility and eventually restricting its recruitment to centrosomes for activation. With this mechanism, BP nanomaterials show great anticancer potential in tumour xenografted mice. Together, our study reveals a molecular mechanism for the tumoricidal properties of BP and proposes a direction for biomedical application of nanomaterials by exploring their intrinsic bioactivities.

A novel pyrrole-imidazole polyamide targets Aurora kinase A and suppresses tumor growth in vivo.

Aurora kinase A (Aurora A) plays a critical role in regulating cell mitotic progression and has been considered as a promising drug target for cancer therapy. To develop a novel molecule targeting Aurora A with high selectivity and efficacy, we designed and synthesized a pyrrole-imidazole polyamide (PIP) Hoechst conjugate, PIP-Ht, targeting to a cell-cycle regulated DNA sequence locating at the promoter of human Aurora A gene (AURKA). PIP-Ht potently suppressed AURKA promoter activities, mRNA expression and protein level, induced tumor cell cycle delay and inhibited tumor cell proliferation in vitro. Furthermore, subcutaneous injection of PIP-Ht into mice bearing human cancer xenografts induced significant tumor growth suppression and cell apoptosis. Collectively, PIP-Ht exhibits the potential as an effective therapeutic candidate for the tumor treatment.

DENV NS1 and MMP-9 cooperate to induce vascular leakage by altering endothelial cell adhesion and tight junction.

Dengue virus (DENV) is a mosquito-borne pathogen that causes a spectrum of diseases including life-threatening dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). Vascular leakage is a common clinical crisis in DHF/DSS patients and highly associated with increased endothelial permeability. The presence of vascular leakage causes hypotension, circulatory failure, and disseminated intravascular coagulation as the disease progresses of DHF/DSS patients, which can lead to the death of patients. However, the mechanisms by which DENV infection caused the vascular leakage are not fully understood. This study reveals a distinct mechanism by which DENV induces endothelial permeability and vascular leakage in human endothelial cells and mice tissues. We initially show that DENV2 promotes the matrix metalloproteinase-9 (MMP-9) expression and secretion in DHF patients' sera, peripheral blood mononuclear cells (PBMCs), and macrophages. This study further reveals that DENV non-structural protein 1 (NS1) induces MMP-9 expression through activating the nuclear factor κB (NF-κB) signaling pathway. Additionally, NS1 facilitates the MMP-9 enzymatic activity, which alters the adhesion and tight junction and vascular leakage in human endothelial cells and mouse tissues. Moreover, NS1 recruits MMP-9 to interact with ß-catenin and Zona occludens protein-1/2 (ZO-1 and ZO-2) and to degrade the important adhesion and tight junction proteins, thereby inducing endothelial hyperpermeability and vascular leakage in human endothelial cells and mouse tissues. Thus, we reveal that DENV NS1 and MMP-9 cooperatively induce vascular leakage by impairing endothelial cell adhesion and tight junction, and suggest that MMP-9 may serve as a potential target for the treatment of hypovolemia in DSS/DHF patients.

Coibamide A kills cancer cells through inhibiting autophagy.

Natural products are useful tools for biological mechanism research and drug discovery. Due to the excellent tumor cell growth inhibitory profile and sub-nanomolar potency, Coibamide A (CA), an N-methyl-stabilized depsipeptide isolated from marine cyanobacterium, has been considered as a promising lead compound for cancer treatment. However, the molecular anti-cancer mechanism of the action of CA remains unclear. Here, we showed that CA treatment induced caspase-independent cell death in breast cancer cells. CA treatment also led to severe lysosome defects, which was ascribed to the impaired glycosylation of lysosome membrane protein LAMP1 and LAMP2. As a consequence, the autophagosome-lysosome fusion was blocked upon CA treatment. In addition, we presented evidence that this autophagy defect partially contributed to the CA treatment-induced tumor cell death. Together, our work uncovers a novel mechanism underlying the anti-cancer action of CA, which will promote its further application for cancer therapy.

One-pot synthesis of VO x /Al 2 O 3 as efficient catalysts for propane dehydrogenation.

Vanadium oxides, as highly efficiently catalysts, are widely applied in various catalytic reactions, such as the dehydrogenation of light alkanes and epoxidation of alkenes. In this paper, a series of VO x /Al 2 O 3 catalysts were fabricated by the 1-pot method for catalytic propane dehydrogenation. The results indicated that the VO x /Al 2 O 3 catalysts with loading of 10 wt.% vanadium exhibited optimized catalytic performance. The as-prepared catalysts were characterized by N 2 adsorption-desorption, XRD, TEM, H 2 -TPR, and XPS to explore the texture properties, morphology, and electronic environment of vanadium. In addition, several vanadium catalysts were also prepared by the incipient wetness impregnation (IWI) method to compare their catalytic performance with the 1-pot synthesized catalysts. The catalysts synthesized by the 1-pot method exhibited higher selectivity of propylene and longer catalyst lifetime at high propane conversion when compared to the counterpart synthesized by the IWI method.

Selfish mutations dysregulating RAS-MAPK signaling are pervasive in aged human testes.

Mosaic mutations present in the germline have important implications for reproductive risk and disease transmission. We previously demonstrated a phenomenon occurring in the male germline, whereby specific mutations arising spontaneously in stem cells (spermatogonia) lead to clonal expansion, resulting in elevated mutation levels in sperm over time. This process, termed "selfish spermatogonial selection," explains the high spontaneous birth prevalence and strong paternal age-effect of disorders such as achondroplasia and Apert, Noonan and Costello syndromes, with direct experimental evidence currently available for specific positions of six genes (FGFR2, FGFR3, RET, PTPN11, HRAS, and KRAS). We present a discovery screen to identify novel mutations and genes showing evidence of positive selection in the male germline, by performing massively parallel simplex PCR using RainDance technology to interrogate mutational hotspots in 67 genes (51.5 kb in total) in 276 biopsies of testes from five men (median age, 83 yr). Following ultradeep sequencing (about 16,000×), development of a low-frequency variant prioritization strategy, and targeted validation, we identified 61 distinct variants present at frequencies as low as 0.06%, including 54 variants not previously directly associated with selfish selection. The majority (80%) of variants identified have previously been implicated in developmental disorders and/or oncogenesis and include mutations in six newly associated genes (BRAF, CBL, MAP2K1, MAP2K2, RAF1, and SOS1), all of which encode components of the RAS-MAPK pathway and activate signaling. Our findings extend the link between mutations dysregulating the RAS-MAPK pathway and selfish selection, and show that the aging male germline is a repository for such deleterious mutations.

Targeting Polo-like Kinase 1 by a Novel Pyrrole-Imidazole Polyamide-Hoechst Conjugate Suppresses Tumor Growth In Vivo.

The serine/threonine kinase Polo-like kinase 1 (Plk1) plays a pivotal role in cell proliferation and has been validated as a promising anticancer drug target. However, very limited success has been achieved in clinical applications using existing Plk1 inhibitors, due to lack of sufficient specificity toward Plk1. To develop a novel Plk1 inhibitor with high selectivity and efficacy, we designed and synthesized a pyrrole-imidazole polyamide-Hoechst conjugate, PIP3, targeted to specific DNA sequence in the PLK1 promoter. PIP3 could specifically inhibit the cell cycle-regulated Plk1 expression and consequently retard tumor cell growth. Cancer cells treated with PIP3 exhibited severe mitotic defects and increased apoptosis, whereas normal cells were not affected by PIP3 treatment. Furthermore, subcutaneous injection of PIP3 into mice bearing human cancer xenografts induced significant tumor growth suppression with low host toxicity. Therefore, PIP3 exhibits the potential as an effective agent for targeted cancer therapy. Mol Cancer Ther; 17(5); 988-1002. ©2018 AACR.

Mammalian Numb protein antagonizes Notch by controlling postendocytic trafficking of the Notch ligand Delta-like 4.

The biological antagonism between the signaling proteins Numb and Notch has been implicated in the regulation of many developmental processes, especially in asymmetric cell division. Mechanistic studies show that Numb inactivates Notch via endocytosis and proteasomal degradation that directly reduce Notch protein levels at the cell surface. However, some aspects of how Numb antagonizes Notch remain unclear. Here, we report a novel mechanism in which Numb acts as a Notch antagonist by controlling the intracellular destination and stability of the Notch ligand Delta-like 4 (Dll4) through a postendocytic-sorting process. We observed that Numb/Numblike knockdown increases the stability and cell-surface accumulation of Dll4. Further study indicated that Numb acts as a sorting switch to control the postendocytic trafficking of Dll4. Of note, the Numb/Numblike knockdown decreased Dll4 delivery to the lysosome, while increasing the recycling of Dll4 to the plasma membrane. Moreover, we demonstrate that this enrichment of Dll4 at the cell surface within Numb/Numblike knockdown cells could activate Notch signaling in neighboring cells. We also provide evidence that Numb negatively controls the Dll4 plasma membrane recycling through a well-documented recycling regulator protein AP1. In conclusion, our study has uncovered a molecular mechanism whereby Numb regulates the endocytic trafficking of the Notch ligand Dll4. Our findings provide a new perspective on how Numb counteracts Notch signaling and sheds additional critical insights into the antagonistic relationship between Numb and Notch signaling.

Numb positively regulates autophagic flux via regulating lysosomal function.

Autophagy is a lysosome-dependent catabolic process involving in the degradation and recycling of unnecessary or damaged proteins and organelles. Emerging evidence indicates that autophagy dysfunction is closely related to various human diseases including cancer, aging, myopathies and neurodegenerative disorders. Here, using genetic knockdown, we uncover the role of Numb, an endocytic adaptor protein, in regulating the late steps of autophagy. We found that Numb depletion led to the accumulation of autophagic vacuole, as verified by RFP-LC3 staining combined with transmission electron microscopy. Further investigation indicated that Numb depletion impaired autophagic degradation through inhibiting the activities of lysosomal enzymes (Cathepsin D, ß-glucuronidase and ß-glucosidase). Moreover, Numb depletion induced elevation of lysosomal pH values and decrease of glycosylated lysosome-associated membrane proteins. We further observed that Rab7 activity was inhibited in Numb-depleted cells. Together, our findings revealed a novel function of Numb and its likely mechanism in regulation of autophagy events.